terminated antigens on the human sperm surface. Fertil Steril 1991;S63. 5. Margalioth EJ, Cooper GW, Taney FR, Scholl GM, Rosenfeld DL. Capacitated sperm cells react with different types of antisperm antibodies than fresh ejaculated sperm. Fertil Steril 1992;57:393-8.
Autoantibodies in In Vitro Fertilization Patients
To the Editor: We read with interest the paper by Fisch et al. (1), in which they suggest an increased level of antiphospholipid antibodies in patients undergoing in vitro fertilization (IVF) in comparison with controls. They conclude that serum levels of antiphospholipid antibodies increase after IVF treatment. We disagree with their interpretation of these data. The study in fact demonstrates that patients who reach IVF have higher phospholipid antibody levels than controls and possibly higher levels than other infertility patients. This is nothing new! In a quite dated publication from our laboratory (2), we reported a statistically highly unusual 40% incidence of autoantibody abnormalities in IVF patients. However, autoantibody positive patients demonstrated even higher autoantibody levels within follicular fluid. Phospholipid antibodies in particular demonstrated a complete abscence of the usual physiological gradient between serum and follicular fluid and appeared in affected females in higher concentrations in follicles than serum of normal controls. This raised the question whether phospholipid antibodies are produced within ovaries. Based on the data by Fisch et al. and our work, it is tempting to speculate that autoantibody problems contribute to the infertility of patients who reach IVF. We have presented supporting evidence for such an effect of autoantibodies in a number of publications (2, 3). These data also suggest that ovarian dysfunction may be associated with the presence of phospholipid antibodies. Premature ovarian failure is now in a majority of cases considered an autoimmune process. This mandates the conclusion that an autoimmune-mediated form of ovarian dysfunction exists that precedes the endstage condition of ovarian failure, which could be expected to be represented in a disproportionate proportion among patients reaching IVF programs. We concur with Fisch et al. that antibodies to phospholipid epitopes appear to playa special role in reproductive failure. They nevertheless only serve Vol. 58, No.4, October 1992
as markers of a broadly based {3 lymphocyte dYRfunction that also involves other autoantibody groupings (2, 3). Their paper contains a number of additional errors. They are incorrect in claiming that lupus anticoagulant activity is always associated with antibodies to phosphatidylserine. To my knowledge, not even the authors of the quoted reference (4) make this claim anymore. One also has to question whether autoantibody levels in fact decline with age. There is considerable data in the literature to make exactly the opposite point. Because we have demonstrated in a variety of ways that abnormal autoantibodies, and especially phospholipid antibodies, are associated with infertility, IVF patients at our center are routinely screened for autoantibody abnormalities. We have consistently found a 40% to 60% incidence of autoantibody abnormalities in prospective IVF patients. We presently are trying to establish a satisfactory treatment option for affected patients that will improve their IVF outcome. Interestingly, Kemeter and Feichtinger (5) reported years ago that corticosteroid therapy improves IVF pregnancy rates.
Norbert Gleicher, M.D. Donna Pratt, M.D. Alan B. Dudkiewicz, Ph.D. The Foundation for Reproductive Medicine and The Center for Human Reproduction Chicago, Illinois December 26, 1992
REFERENCES 1. Fisch B, Rikover Y, Shohat L, Zurgil N, Tadir Y, Ovadia J,
2.
3.
4.
5.
et al. The relationship between in vitro fertilization and naturally occurring antibodies: evidence for increased production of antiphospholipid autoantibodies. Fertil Steril 1991;56: 718-24. El-Roeiy A, Gleicher N, Friberg J, Contino E, Dudkiewicz A. Correlation between peripheral blood and follicular fluid autoantibodies and impact on in vitro fertilization. Obstet Gynecol 1987;70:163-9. Gleicher N, El-Roeiy A, Contino E, Friberg J. Reproductive failure because of autoantibodies: unexplained infertility and pregnancy wastage. Am J Obstet Gynecol1989;160:1376-85. Ware Branch D, Rote NS, Dostal DA, Scott JR. Association of lupus anticoagulant with antibody against phosphatidylserine. Clin Immunol Immunopathol 1987;42:663-8. Kemeter P, Feichtinger W. Prednisolon verbessert die Schwangerschaftsrate der IVF. Eine prospective randomisierte Studie. Fertilitat 1986;2:71-6.
Letters-to-the-editor
863
Reply of the Authors: Thank you for offering us the opportunity to comment on the interesting letter by Gleicher et al. They have taken issue with the interpretation of the data presented in our paper (1). The aim of our study was to assess the effect of the extreme hormonal fluctuations occurring during in vitro fertilization (IVF) treatment on autoantibody levels in women who have already undergone at least one IVF cycle. Thus, these data, and subsequently the conclusions drawn, neither contradict nor support the hypothesis suggested by Gleicher et al. that "patients who reach IVF have higher phospholipid antibody levels." A longitudinal study currently being undertaken in our laboratory on serum samples obtained before, during, and after the first IVF treatment cycle of each individual patient addresses this specific question. Moreover, in our opinion, the data presented by these authors (2) do not necessarily concur with their own theory either: they obtained the sera from IVF patients at one time point only on the day of oocyte retrieval, i.e., after hormonal treatment. Furthermore, they compared between seropositive and seronegative patients within the same group, without including normal controls in their study. The remark concerning the correlation between ovarian dysfunction and antiphospholipid antibody levels may be important but is irrelevant to our study. As described in the Materials and Methods section of our article (1), all patients included were ovulating regularly, had normal endocrine profile, and responded well to superovulation protocols, thus excluding occult ovarian failure in our study group. Regarding the suggested association between lupus anticoagulant activity and antibodies to phosphatidylserine, we were not claiming, as stated in the letter, but rather referring to and faithfully quoting from the article by Ware Branch et ai. (3) that "lupus anticoagulant activity is always associated with the presence of autoantibodies against phosphatidylserine." Even in a later article by these researchers (4) (unavailable to us at the time of preparation of our manuscript), they emphasized the significant association between antibodies to phosphatidylserine and lupus anticoagulant, although not as firmly as in their previous paper quoted by us ". . . antiphosphatidylserine assay correlates best, although not totally, with the presence of lupus anticoagulant and that the antiphosphatidylserine assay is more sensitive than testing for anticardiolipin" (4).
864
Letters-to-the-editor
Because we could not show any correlation of our results to the patients' age (and, therefore, even avoided presenting these data), we find it very peculiar that Gleicher and colleagues argue with us on this matter. Indeed, we should have referred to the article by EI-Roeiy et al. (2); however, this work differs significantly from ours in several major points. [1] We did not subdivide our cases to "autoantibody positive" and "autoantibody negative" study groups but rather compared serum activity of patients undergoing IVF to proper controls. [2] Our control group consisted only of women with no known infertility problems. Obviously, it did not include any IVF patient, in contrast to the work by EI-Roeiy et al. (2). [3] According to our study objective and unlike EIRoeiy et aI., we actually measured changes in antiphospholipid autoantibody levels at three time points along the IVF treatment cycle and presented the data accordingly. Finally, the limited data available to date concerning various aspects of autoantibody levels in infertile women justify further studies. This could shed more light on an interesting subject that may have significant clinical implications. We feel that any well-designed research project will contribute to the accumulating knowledge in this field.
Benjamin Fisch, M.D., Ph.D. Yigal Rikover, M.D. Yona Tadir, M.D. Jardena Ovadia, M.D. Infertility and In Vitro Fertilization Unit Department of Obstetrics and Gynecology Beilinson Medical Center Sackler School of Medicine, Tel-Aviv University Petah Tikva, Israel !lana Yron, Ph.D. Lea Shohat, M.Sc. Isaac P. Witz, Ph.D. Department of Microbiology-Cell Biology and the Moise and Frida Eskenasy Institute for Cancer Research The George S. Wise Faculty of Life Sciences Tel-Aviv University Tel-Aviv, Israel Neomi Zurgil, Ph.D. BioHyTec Ramat-Gan, Israel June 9, 1992
Fertility and Sterility
REFERENCES 1. Fisch B, Rikover Y, Shohat L, Zurgil N, Tadir Y, Ovadia J,
et al. The relationship between in vitro fertilization and naturally occurring antibodies: evidence for increased production of antiphospholipid autoantibodies. Fertil Steril 1991;56:71824. 2. El-Roeiy A, Gleicher N, Friberg J, Confino E, Dudkiewicz A. Correlation between peripheral blood and follicular fluid autoantibodies and impact on in vitro fertilization. Obstet GynecoI1987;70:163-70. 3. Ware Branch D, Rote NS, Dostal DA, Scott JR. Association of lupus anticoagulant with antibody against phosphatidylserine. Clin Immunol Immunopathol 1987;42:63-75. 4. Rote NS, Dostal-Johnson D, Ware Branch D. Antiphospholipid antibodies and recurrent pregnancy loss: correlation between the activated partial thromboplastin time and antibodies against phosphatidylserine and cardiolipin. Am J Obstet GynecoI1990;163:575-84.
Protection Against Antisperm Antibodies?
To the Editors: Involved in ongoing studies of the lymphoreticular system of the male accessory sexual glands of reproduction, particularly the prostate, and their contributory role in health and diseases of these tissues, we read with interest the report by Bukovsky et ai. (1) of the identification of soluble Fc-gamma receptor III (Fc"), RIll) in the male accessory sexual glands. Having earlier reported the presence of Fc")' RIll in the human prostate, we thought brief comment of these initial and subsequent observations of soluble FC")'RIII and some of the implications of their immunoregulatory effect in the human immunodeficiency virus (HIV) infection and the maternal-fetal relationship would be of interest. Initially, single and double immunofluorescence (IF) with an fluorescein isothiocyanate-conjugated anti-human monoclonal antibody to Leu-lla (CD16) and a polyclonal anti-human prostate specific antigen (as an epithelial-cell marker) or an antiLeu-M5 (CDllc, a monocyte/macrophage marker for distinguishing stromal IF) with a Texas Red conjugated anti-immunoglobulin were used in evaluating the human prostate. Such studies that disclosed intense focal localization to cells and elements within the fibromuscular stroma were identifiable as histiocytes, mast cells, and collagenase fibers. Contrary to the subsequent studies by Bukovsky et aI., acinar epithelial cells were devoid of staining for FC")'RIII. In subsequent studies, dot-immunoblot analysis of prostate tissue extracts and seminal Vol. 58, No.4, October 1992
plasma with an anti-human monoclonal antibody to Leu-lla permitted the identification of soluble FC")'RIII. The localization of Fc"), RIll in the prostate by Bukovsky et ai. and ourselves has suggested a possible prostatic origin for soluble Fc")' RIll in seminal plasma. This suggestion is further strengthened by observations of an increase in Fc")' RIll positive peripheral blood lymphocytes after their incubation in prostatic but not seminal vesicle fluids (2). The biological functions of soluble FC")' RIll remain somewhat speculative. Nonetheless, two areas of current interest where the immunoregulatory effect of soluble FC")' RIll and their modulation may play pivotal roles are in HIV infection and the maternal-fetal relationship. In a recent study of serum of HIV infected patients (3), a dramatically significant inverse correlation between the level of soluble FC")' RIll with progression of disease to clinically acquired immune deficiency syndrome (AIDS) has been noted. A further significant correlation was observed between changes of Fc")' RIll levels and a decrease in CD4+ cells, increase in p24 antigen, and decrease in antip24 antibody, the three most important (to date) biological and clinical prognostic parameters for AIDS (3). These observations, together with those demonstrating that FC")' RIll mediates uptake of HIV enhancing antibody complexes into human macrophages (4), further implicates the involvement of Fc")'RIII in the course of HIV disease. In looking at a second current and possibly pivotal role of the immunoregulatory effect of soluble FC")'RIII, the earlier localization ofFC")'R to the placenta (5 and references therein) becomes of perhaps particular further biological significance to regulation of the maternal-fetal immune response in fertilization and pregnancy. Noteworthy herein may lie perhaps a unique evolutionary mechanism to ensure successful implantation. In this regard, seminal plasma, traditionally thought of primarily as a vehicle for the transport of spermatozoa as a means of perpetuating the species via insemination and fertilization, may in view of its content of Fc")' RIll in contributing, in part, to its known IR activity provide a unique homeostatic mechanism regulating the maternal-fetal immune response.
Terry C. Whyard, M.A. Department of Urology State University of New York at Stony Brook School of Medicine Stony Brook, New York Letters-to-the-editor
865
Autoantibodies in In Vitro Fertilization Patients
To the Editor: We read with interest the paper by Fisch et al. (1), in which they suggest an increased level of antiphospholipid antibodies in patients undergoing in vitro fertilization (IVF) in comparison with controls. They conclude that serum levels of antiphospholipid antibodies increase after IVF treatment. We disagree with their interpretation of these data. The study in fact demonstrates that patients who reach IVF have higher phospholipid antibody levels than controls and possibly higher levels than other infertility patients. This is nothing new! In a quite dated publication from our laboratory (2), we reported a statistically highly unusual 40% incidence of autoantibody abnormalities in IVF patients. However, autoantibody positive patients demonstrated even higher autoantibody levels within follicular fluid. Phospholipid antibodies in particular demonstrated a complete abscence of the usual physiological gradient between serum and follicular fluid and appeared in affected females in higher concentrations in follicles than serum of normal controls. This raised the question whether phospholipid antibodies are produced within ovaries. Based on the data by Fisch et al. and our work, it is tempting to speculate that autoantibody problems contribute to the infertility of patients who reach IVF. We have presented supporting evidence for such an effect of autoantibodies in a number of publications (2, 3). These data also suggest that ovarian dysfunction may be associated with the presence of phospholipid antibodies. Premature ovarian failure is now in a majority of cases considered an autoimmune process. This mandates the conclusion that an autoimmune-mediated form of ovarian dysfunction exists that precedes the endstage condition of ovarian failure, which could be expected to be represented in a disproportionate proportion among patients reaching IVF programs. We concur with Fisch et al. that antibodies to phospholipid epitopes appear to playa special role in reproductive failure. They nevertheless only serve Vol. 58, No.4, October 1992
as markers of a broadly based {3 lymphocyte dYRfunction that also involves other autoantibody groupings (2, 3). Their paper contains a number of additional errors. They are incorrect in claiming that lupus anticoagulant activity is always associated with antibodies to phosphatidylserine. To my knowledge, not even the authors of the quoted reference (4) make this claim anymore. One also has to question whether autoantibody levels in fact decline with age. There is considerable data in the literature to make exactly the opposite point. Because we have demonstrated in a variety of ways that abnormal autoantibodies, and especially phospholipid antibodies, are associated with infertility, IVF patients at our center are routinely screened for autoantibody abnormalities. We have consistently found a 40% to 60% incidence of autoantibody abnormalities in prospective IVF patients. We presently are trying to establish a satisfactory treatment option for affected patients that will improve their IVF outcome. Interestingly, Kemeter and Feichtinger (5) reported years ago that corticosteroid therapy improves IVF pregnancy rates.
Norbert Gleicher, M.D. Donna Pratt, M.D. Alan B. Dudkiewicz, Ph.D. The Foundation for Reproductive Medicine and The Center for Human Reproduction Chicago, Illinois December 26, 1992
REFERENCES 1. Fisch B, Rikover Y, Shohat L, Zurgil N, Tadir Y, Ovadia J,
2.
3.
4.
5.
et al. The relationship between in vitro fertilization and naturally occurring antibodies: evidence for increased production of antiphospholipid autoantibodies. Fertil Steril 1991;56: 718-24. El-Roeiy A, Gleicher N, Friberg J, Contino E, Dudkiewicz A. Correlation between peripheral blood and follicular fluid autoantibodies and impact on in vitro fertilization. Obstet Gynecol 1987;70:163-9. Gleicher N, El-Roeiy A, Contino E, Friberg J. Reproductive failure because of autoantibodies: unexplained infertility and pregnancy wastage. Am J Obstet Gynecol1989;160:1376-85. Ware Branch D, Rote NS, Dostal DA, Scott JR. Association of lupus anticoagulant with antibody against phosphatidylserine. Clin Immunol Immunopathol 1987;42:663-8. Kemeter P, Feichtinger W. Prednisolon verbessert die Schwangerschaftsrate der IVF. Eine prospective randomisierte Studie. Fertilitat 1986;2:71-6.
Letters-to-the-editor
863
Reply of the Authors: Thank you for offering us the opportunity to comment on the interesting letter by Gleicher et al. They have taken issue with the interpretation of the data presented in our paper (1). The aim of our study was to assess the effect of the extreme hormonal fluctuations occurring during in vitro fertilization (IVF) treatment on autoantibody levels in women who have already undergone at least one IVF cycle. Thus, these data, and subsequently the conclusions drawn, neither contradict nor support the hypothesis suggested by Gleicher et al. that "patients who reach IVF have higher phospholipid antibody levels." A longitudinal study currently being undertaken in our laboratory on serum samples obtained before, during, and after the first IVF treatment cycle of each individual patient addresses this specific question. Moreover, in our opinion, the data presented by these authors (2) do not necessarily concur with their own theory either: they obtained the sera from IVF patients at one time point only on the day of oocyte retrieval, i.e., after hormonal treatment. Furthermore, they compared between seropositive and seronegative patients within the same group, without including normal controls in their study. The remark concerning the correlation between ovarian dysfunction and antiphospholipid antibody levels may be important but is irrelevant to our study. As described in the Materials and Methods section of our article (1), all patients included were ovulating regularly, had normal endocrine profile, and responded well to superovulation protocols, thus excluding occult ovarian failure in our study group. Regarding the suggested association between lupus anticoagulant activity and antibodies to phosphatidylserine, we were not claiming, as stated in the letter, but rather referring to and faithfully quoting from the article by Ware Branch et ai. (3) that "lupus anticoagulant activity is always associated with the presence of autoantibodies against phosphatidylserine." Even in a later article by these researchers (4) (unavailable to us at the time of preparation of our manuscript), they emphasized the significant association between antibodies to phosphatidylserine and lupus anticoagulant, although not as firmly as in their previous paper quoted by us ". . . antiphosphatidylserine assay correlates best, although not totally, with the presence of lupus anticoagulant and that the antiphosphatidylserine assay is more sensitive than testing for anticardiolipin" (4).
864
Letters-to-the-editor
Because we could not show any correlation of our results to the patients' age (and, therefore, even avoided presenting these data), we find it very peculiar that Gleicher and colleagues argue with us on this matter. Indeed, we should have referred to the article by EI-Roeiy et al. (2); however, this work differs significantly from ours in several major points. [1] We did not subdivide our cases to "autoantibody positive" and "autoantibody negative" study groups but rather compared serum activity of patients undergoing IVF to proper controls. [2] Our control group consisted only of women with no known infertility problems. Obviously, it did not include any IVF patient, in contrast to the work by EI-Roeiy et al. (2). [3] According to our study objective and unlike EIRoeiy et aI., we actually measured changes in antiphospholipid autoantibody levels at three time points along the IVF treatment cycle and presented the data accordingly. Finally, the limited data available to date concerning various aspects of autoantibody levels in infertile women justify further studies. This could shed more light on an interesting subject that may have significant clinical implications. We feel that any well-designed research project will contribute to the accumulating knowledge in this field.
Benjamin Fisch, M.D., Ph.D. Yigal Rikover, M.D. Yona Tadir, M.D. Jardena Ovadia, M.D. Infertility and In Vitro Fertilization Unit Department of Obstetrics and Gynecology Beilinson Medical Center Sackler School of Medicine, Tel-Aviv University Petah Tikva, Israel !lana Yron, Ph.D. Lea Shohat, M.Sc. Isaac P. Witz, Ph.D. Department of Microbiology-Cell Biology and the Moise and Frida Eskenasy Institute for Cancer Research The George S. Wise Faculty of Life Sciences Tel-Aviv University Tel-Aviv, Israel Neomi Zurgil, Ph.D. BioHyTec Ramat-Gan, Israel June 9, 1992
Fertility and Sterility
REFERENCES 1. Fisch B, Rikover Y, Shohat L, Zurgil N, Tadir Y, Ovadia J,
et al. The relationship between in vitro fertilization and naturally occurring antibodies: evidence for increased production of antiphospholipid autoantibodies. Fertil Steril 1991;56:71824. 2. El-Roeiy A, Gleicher N, Friberg J, Confino E, Dudkiewicz A. Correlation between peripheral blood and follicular fluid autoantibodies and impact on in vitro fertilization. Obstet GynecoI1987;70:163-70. 3. Ware Branch D, Rote NS, Dostal DA, Scott JR. Association of lupus anticoagulant with antibody against phosphatidylserine. Clin Immunol Immunopathol 1987;42:63-75. 4. Rote NS, Dostal-Johnson D, Ware Branch D. Antiphospholipid antibodies and recurrent pregnancy loss: correlation between the activated partial thromboplastin time and antibodies against phosphatidylserine and cardiolipin. Am J Obstet GynecoI1990;163:575-84.
Protection Against Antisperm Antibodies?
To the Editors: Involved in ongoing studies of the lymphoreticular system of the male accessory sexual glands of reproduction, particularly the prostate, and their contributory role in health and diseases of these tissues, we read with interest the report by Bukovsky et ai. (1) of the identification of soluble Fc-gamma receptor III (Fc"), RIll) in the male accessory sexual glands. Having earlier reported the presence of Fc")' RIll in the human prostate, we thought brief comment of these initial and subsequent observations of soluble FC")'RIII and some of the implications of their immunoregulatory effect in the human immunodeficiency virus (HIV) infection and the maternal-fetal relationship would be of interest. Initially, single and double immunofluorescence (IF) with an fluorescein isothiocyanate-conjugated anti-human monoclonal antibody to Leu-lla (CD16) and a polyclonal anti-human prostate specific antigen (as an epithelial-cell marker) or an antiLeu-M5 (CDllc, a monocyte/macrophage marker for distinguishing stromal IF) with a Texas Red conjugated anti-immunoglobulin were used in evaluating the human prostate. Such studies that disclosed intense focal localization to cells and elements within the fibromuscular stroma were identifiable as histiocytes, mast cells, and collagenase fibers. Contrary to the subsequent studies by Bukovsky et aI., acinar epithelial cells were devoid of staining for FC")'RIII. In subsequent studies, dot-immunoblot analysis of prostate tissue extracts and seminal Vol. 58, No.4, October 1992
plasma with an anti-human monoclonal antibody to Leu-lla permitted the identification of soluble FC")'RIII. The localization of Fc"), RIll in the prostate by Bukovsky et ai. and ourselves has suggested a possible prostatic origin for soluble Fc")' RIll in seminal plasma. This suggestion is further strengthened by observations of an increase in Fc")' RIll positive peripheral blood lymphocytes after their incubation in prostatic but not seminal vesicle fluids (2). The biological functions of soluble FC")' RIll remain somewhat speculative. Nonetheless, two areas of current interest where the immunoregulatory effect of soluble FC")' RIll and their modulation may play pivotal roles are in HIV infection and the maternal-fetal relationship. In a recent study of serum of HIV infected patients (3), a dramatically significant inverse correlation between the level of soluble FC")' RIll with progression of disease to clinically acquired immune deficiency syndrome (AIDS) has been noted. A further significant correlation was observed between changes of Fc")' RIll levels and a decrease in CD4+ cells, increase in p24 antigen, and decrease in antip24 antibody, the three most important (to date) biological and clinical prognostic parameters for AIDS (3). These observations, together with those demonstrating that FC")' RIll mediates uptake of HIV enhancing antibody complexes into human macrophages (4), further implicates the involvement of Fc")'RIII in the course of HIV disease. In looking at a second current and possibly pivotal role of the immunoregulatory effect of soluble FC")'RIII, the earlier localization ofFC")'R to the placenta (5 and references therein) becomes of perhaps particular further biological significance to regulation of the maternal-fetal immune response in fertilization and pregnancy. Noteworthy herein may lie perhaps a unique evolutionary mechanism to ensure successful implantation. In this regard, seminal plasma, traditionally thought of primarily as a vehicle for the transport of spermatozoa as a means of perpetuating the species via insemination and fertilization, may in view of its content of Fc")' RIll in contributing, in part, to its known IR activity provide a unique homeostatic mechanism regulating the maternal-fetal immune response.
Terry C. Whyard, M.A. Department of Urology State University of New York at Stony Brook School of Medicine Stony Brook, New York Letters-to-the-editor
865
